The detection of Plasmodium falciparum in human saliva samples
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Date
2011
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Abstract
Malaria still persists as a major public health scourge, claiming a global toll of up to 500 million clinical cases, mostly on African children. Current effective malaria diagnostic methods necessitate blood withdrawal from patients for accurate P. falciparum identification. Risks introduced by blood withdrawal can causes communities to be less cooperative to donate blood during malaria surveillance studies. Previous studies showed that saliva could be used to detect malaria by molecular techniques. This study, therefore, sought to establish the constituents of human saliva that harbours parasite DNA in malaria infected subjects. Furthermore the study optimised the use of Saliva as an alternative malaria DNA source in malaria infected patients. A total of 88 subjects were enrolled in this study, 35 (40, 7 %) were males and 51 (59, 3 %) were females. The age range was from 3 months to 99 years old (mean = 29.6 years; median = 18 years). Blood was drawn from each subject for subsequent use in microscopic examination and PCR tests. Saliva samples were also collected for PCR on Saliva derived parasite DNA. DNA was extracted using commercial kit on different saliva fractions from 46 malaria positive (thick film positive & blood PCR positives) individuals and 45 malaria negative individuals. Nested PCR was used to amplify malaria the Plasmodium falciparum dihydrofolate reductase (pfdhfr) gene. Generally PCR conducted on DNA purified from both blood and saliva was more sensitive than conventional microscopy, as previously reported. The pellet fraction of saliva was a more reliable and sensitive source of amplifiable parasite DNA compared to the soluble saliva fraction. After PCR optimisation amplification was enhanced 94.1% (sensitivity) and 97% (specificity) against DNA derived from blood as the gold standard. This study further confirms that saliva samples are a reliable non-invasive alternative to blood for the PCR detection of malaria, as previously reported. The source of DNA was primarily in the pellet fraction though; however, refinement is still needed to identify the exact source of DNA in that fraction.
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Dissertation submitted to the department of Biochemistry and Microbiology, Faculty of Science and Agriculture, University of Zululand in partial fulfilment of the requirements for the Masters (MSc) degree in Biochemistry, 2011.